Apparent intracellular Mg2+ buffering in neurons of the leech Hirudo medicinalis

The apparent intracellular Mg2+ buffering, or muffling (sum of processes that damp changes in the free intracellular Mg2+ concentration, [Mg2+](i), e.g., buffering, extrusion, and sequestration), was investigated in Retzius neurons of the leech Hirudo medicinalis by iontophoretic injection of H+, OH-, or Mg2+. Simultaneously, changes in intracellular pH and the intracellular Mg2+, Na+, or K+ concentration were recorded with triple-barreled ion-selective microelectrodes. Cell volume changes were monitored measuring the tetramethylammonium (TMA) concentration in TMA-loaded neurons. Control measurements were carried out in electrolyte droplets (diameter 100-200 microm) placed on a silver wire under paraffin oil. Droplets with or without ATP, the presumed major intracellular Mg2+ buffer, were used to quantify the pH dependence of Mg2+ buffering and to determine the transport index of Mg2+ during iontophoretic injection. The observed pH dependence of [Mg2+](i) corresponded to what would be expected from Mg2+ buffering through ATP. The quantity of Mg2+ muffling, however, was considerably larger than what would be expected if ATP were the sole Mg2+ buffer. From the decrease in Mg2+ muffling in the nominal absence of extracellular Na+ it was estimated that almost 50% of the ATP-independent muffling is due to the action of Na+/Mg2+ antiport.